This invention relates generally to a process for the preparation of immunogenic detoxified polysaccharide-outer membrane protein complexes from bacteria and more particularly to the use of said complexes, as vaccines, to protect animals against infections by the bacteria from which it has been derived comprising administering to an animal a pharmaceutical composition containing the detoxified polysaccharide-protein complexes.
The virulence of certain gram-negative bacteria is enhanced by the presence of a capsule which envelopes the outer membrane and is made up of different components among which are polysaccharides and lipopolysaccharides.
The outer membrane proteins of gram-negative bacteria such as Haemophilus influenza type b, Neisseria gonorrhoeae, Escherichia coli, Pseudomonas aeruginosa, and Neisseria meningitidis have been shown to induce bactericidal antibodies in man, both when encountered in the course of natural infections and when given as a vaccine in which the proteins are noncovalently complexed to the capsular polysaccharide. Vaccines of this type, as exemplified by Neisseria meningitidis, have been studied by several groups to determine their potential for providing protection against group B meningococcal disease. In most instances the outer membrane proteins from a serotype 2a strain have been used together with group B polysaccharide. Although vaccines of this type have shown promise in terms of safety and immunogenicity, there are a number of problems which need to be resolved.
When presented as a complex with the outer membrane proteins, the B polysaccharide induces a transient IgM antibody response. These antibodies are bactericidal with rabbit complement, but have little if any bactericidal activity with human complement. This fact together with reports that antibodies to group B polysaccharide cross-react with human fetal and neonatal brain antigens suggest to applicants that these antibodies are probably not protective and an alternative to group B polysaccharide is needed in future vaccine preparations.
A second problem is the great antigenic diversity and variability of the outer membrane proteins. Meningococci possess multiple outer membrane proteins which are known to vary antigenically from strain to strain. According to the classification scheme of Tsai, et al. described in the Journal of Bacteriology, Volume 146, pages 69 to 78, 1981, 5 major classes of outer membrane proteins are recognized. Of these, classes 1, 2, 3, and 5 have been shown to vary antigenically from strain to strain. In addition, lipopolysaccharides are present and exhibit antigenic variability. From the point of view of vaccine development this creates problems both in terms of vaccine formulation and evaluation of the antibody response to vaccination. The number of different serotype proteins that can safely be included in a single vaccine may be limited by reactogenicity resulting from the residual lipopolysaccharide (LPS) associated with them and probably also by reactogenicity intrinsic to the proteins themselves.
Applicants have evaluated the human bactericidal antibody response to serotype 2b and serotype 15 outer membrane proteins, prepared these proteins relatively free of LPS (less than 1%), and discovered the immunogenicity and safety of these proteins when combined in a single vaccine and solubilized by the tetravalent mixture of A, C, Y, and W135 polysaccharides.
The existence of other methods for preparing vaccines useful against infections caused by gram-negative bacteria are disclosed in U.S. Pat. Nos. 4,451,446; 3,636,192; 3,859,434; 4,356,170; 4,123,520 and 3,978,209 which are hereby incorporated herein by reference.
Although partially effective vaccines are available for treatment of bacterial infections, most vaccines presently known to be employed have limited effectiveness.